Chromatin preparation guidelines and tips

The epigenetic services team at Active Motif has been evolving & optimizing our ChIP chromatin preparation protocols for 10 years. Most sample types are amenable to a standard chromatin preparation protocol but others can be difficult and require a specialized approach.

Below are tips and information about chromatin preparation for various sample types.

Starting amount: 100 to 200 mg as a starting point.
Yield will vary between different tissue types.Protocol: Specialized protocols for fixation and chromatin
preparation.Notes: Requires tissue dissociation using a hand held
homogenizer.

Active Motif has performed ChIP-Seq from >25 different mouse tissues

Mouse tissues used for chromatin preparation

Atria

Brown adipose

Cerebellum

Colon

Diaphragm

Face

Heart

Cerebellum

Hindbrain

Hippocampus

Kidney

White adipose

Lung

Mammary

Muscle

Pancreas

Pituitary

Retina

Thymus

Spinal cord

Spleen

Stomach

Testicle

Sciatic nerve

Tooth bud

Uterus

Liver

Whole brain

Human biopsies and postmortem tissue

Starting amount: 100 to 200 mg as a starting point.
Yield will vary between different tissue types.Protocol: Specialized protocols for fixation and
chromatin preparation.Notes: The protocol is the same as that used for mouse
tissues. This protocol works with all solid tumors
and postmortem tissues.

T cells & B cells

Starting amount: >20 million cellsProtocol: Specialized protocols for fixation and
chromatin preparation.Notes: Requires 2X sonication compared to a standard
preparation. Cell loss can occur during wash steps
after fixation. Yields will be much lower than cell line
preparations using comparable cell numbers.

Mammary tissue & Adipose cells

Starting amount: >500mgProtocol: Specialized protocol for cells containing fat.Notes: Fat content creates layers after spinning lysed
cells and must be separated in order to isolate the
fat cells. More starting material is required than
that used for other tissue types.

Oligodendrocytes and neurons

Starting amount: >5 million cellsProtocol: Specialized protocol that is similar to our
T and B cell protocol.

Drosophila

Starting amount: Varies depending on the tissue type.Protocol: Specialized protocol for Drosophila.Notes: Trichomes, bristles and other structures are not
lysed in the protocol and must be removed during
chromatin preparation.

Starting amount: Varies depending on the species and
tissue type.Protocol: Specialized fixation and chromatin preparation.Notes: Fixation must occur under vacuum, then freezing
with liquid nitrogen followed by grinding the sample
into a powder to facilitate chromatin extraction.